In Vitro and In Vivo Studies of Metabolic Activation of Marrubiin, a Bioactive Constituent from Marrubium Vulgare.

Marrubiin, a furanoid compound, is a well-known diterpenoid lactone isolated from Marrubium vulgare, which displays a wide spectrum of pharmacological effects and potential hepatotoxicity. Considering that marrubiin contains a structural alert, furan ring, metabolic activation may be one of the major metabolic pathways, and the reactive metabolite may be involved in the hepatotoxicity. The present study was carried out to investigate the bioactivation mechanism of marrubiin in rats and humans. Marrubiin was initially metabolized into cis-butene-1,4-dial intermediate, which was readily trapped by glutathione (GSH) and N-acetyl-lysine (NAL) in the microsomal incubations supplemented with NADPH. A total of nine conjugates were detected and identified by high-resolution mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy. M1-M3 and M6 and M7 were characterized as mono-GSH conjugates, and M4 and M5 were identified as bis-GSH conjugates. M8 and M9 were identified as NAL conjugates. In rat bile, five GSH conjugates (M1-M3; M6 and M7) were detected. M1, M8, and M9 were chemically synthesized, and their structures were characterized by 13C NMR. Sulfaphenazole, ticlopidine, and ketoconazole displayed significant inhibitory effect on the bioactivation of marrubiin. Further phenotyping revealed that CYP2C9, CYP2C19, and CYP3A4 were the primary enzymes catalyzing the bioactivation of marrubiin. The current study provides evidence for the CYP-dominated bioactivation of marrubiin to the corresponding cis-butene-1,4-dial intermediate, which enables us to better understand the potential side effects caused by marrubiin.

Silibinin suppresses epithelial-mesenchymal transition in human non-small cell lung cancer cells by restraining RHBDD1.

BACKGROUND: Rhomboid domain containing 1 (RHBDD1) plays a crucial role in tumorigenesis. Silibinin, which is a natural extract from milk thistle, has shown anti-tumor effects against various tumors. Here, we investigate whether silibinin affects the function of RHBDD1 in non-small cell lung cancer (NSCLC) cell proliferation, migration and invasion. METHODS: The Oncomine database and an immunohistochemistry (IHC) assay were used to determine the RHBDD1 expression levels in lung cancer tissues. The associations between RHBDD1 and overall survival rate or clinicopathological parameters were respectively assessed using the Kaplan-Meier overall survival analysis or Chi-squared test. CCK-8 and Transwell assays were applied to analyze cell proliferation, migration and invasion. A549 cells were incubated with increasing concentrations of silibinin. RHBDD1 knockdown and overexpression were achieved via transfection with si-RHBDD1 or RHBDD1 overexpression plasmid, respectively. Western blotting was performed to measure the expressions of epithelial-mesenchymal transition (EMT) markers. RESULTS: We found that overexpression of RHBDD1 in lung cancer tissues correlates with a poor prognosis of survival. Clinical specimen analysis showed that upregulation of RHBDD1 correlates remarkably well with TNM stage and lymph node metastasis. Silibinin suppresses A549 cell proliferation, migration, invasion and EMT in a dose-dependent manner. Importantly, RHBDD1 was downregulated in silibinin-treated A549 cells. RHBDD1 overexpression reversed the suppressive effects of silibinin on A549 cell proliferation, migration, invasion and EMT expression, while its knockdown enhanced them. CONCLUSIONS: These findings shown an anti-tumor impact of silibinin on NSCLC cells via repression of RHBDD1.